Antigen recognition by T lymphocytes is mediated by diverse cell-surface glycoproteins known as T-cell receptors (TCRs) which are composed of alpha and beta (or gamma and delta) chains. In addition to antigenic peptides bound to MHC, alpha/beta TCRs interact with glycolipids presented by CD1 and with a class of disease-causing major subset of human gamma/delta T-cells (V-gamma-2/V-delta-2) recognizes small, aliphatic monoalkyl phosphate antigens from Mycobacterium. Besides conventional alpha/beta and gamma/delta T-cells, the T-cell compartment includes natural killer (NK) T-cells. NK T-cells express NK inhibitory receptors that mediate self-recognition by binding MHC class I. These cells also express TCRs that recognize ceramide-containing glycolipids presented by CD1. Our aim is to elucidate the structural basis for three key aspects of antigen recognition by alpha/beta, gamma/delta and NK T-cells: Determination of the crystal structures of representative TCR beta chain-Sag and SAG-MHC complexes in order to precisely define the diverse strategies that bacterial SAGs have evolved for binding TCR and MHC molecules. The investigator previously determined the structures of a mouse TCR beta chain (VB8.2) complexed with staphylococcal enterotoxins B and C3 (SEB, SEC3). We will now determine the structures of a human TCR beta chain (VB2.1) complexed with toxic shock syndrome toxin-1 (TSST-1) and streptococcal pyrogenic exotoxin C (SPEC). We will also determine the structure of SEC3 complexed with HLA-DR1. This study will then be extended to SEA, SED and SPEC which, unlike SEC3, cross-link MHC molecules on APCs. Determination of the structural basis for antigen recognition by human V-gamma-2/V-delta-2 TCRs. We have recently solved the structure of the TCR V delta domain and shown that it incorporates key structural elements of both antibody and alpha/beta TCR V regions. We will now extend these studies to an associated V-gamma-2/V-delta-2 TCR reactive with non-peptide prenyl pyrophosphates. 3. Determination of the structural basis for self-MHC and CD1 recognition by NK T-cells. We have crystallized a complex between the NK inhibitory receptor Ly-49A and its MHC class I ligand and a structure determination is underway. We will also express soluble forms of a canonical V alpha 14-J alpha 281/V beta 8.2 NK TCR for use in direct binding and co-crystallization experiments with specific CD1/glycolipid antigens.